The goal of this proposal is to elucidate the serotonin signaling pathways through genetic dissection of genes and functional pathways regulated by Selective Serotonin Reuptake Inhibitors (SSRIs). Dysregulation of the serotonergic system has been implicated in a wide-spectrum of neurodevelopmental and psychiatric disorders, and SSRIs have made a major impact on many symptoms of these diseases. SSRIs are thought to exert therapeutic effects by inhibiting the serotonin reuptake transporter (SERT), thereby increasing serotonin signaling, leading to our central hypothesis that genetic dissection of SSRIs-resistant genes will identify not only the genes directly binding the drugs, but also the genes and functional pathways regulated by the drugs, and therefore, will shed lights on the fundamental biology of serotonin signaling and its role in the diseases. Taking the advantage of the availability of serotonin-deficient C. elegans, and the unique ability to monitor gene expression and synapse structure in identified cells in the living worm, this proposal uses direct forward genetic approaches in C. elegans to identify SSRIs-resistant genes in a whole animal. This proposal is based on our preliminary data indicating that the SSRI fluoxetine (Prozac ) targets serotonin postsynaptic targets to regulate the activity of acetylcholine, GABA, and glutamate neurotransmission. In four specific aims, proposed experiments will search genome-wide for mutant worms that are resistant to fluoxetine to elucidate the genetic interactions between SSRIs, serotonin, and the other neurotransmitters. The first aim is to identify fluoxetine- resistant genes. We have established a behavioral paradigm for genetic screening for fluoxetine-resistant mutants. Using this paradigm, we have already identified 5 genes known to be involved in serotonin signaling. In addition, we have isolated 35 new fluoxetine-resistant mutants through an unbiased mutagenesis screen. We will characterize the new mutants and clone the mutant genes. The second aim is to characterize the role of new fluoxetine-resistant genes in regulating acetylcholine, GABA and glutamate neurotransmission, and to organize the drug-resistant genes in functional pathways that couple serotonin and individual neurotransmitter signaling pathways. The third aim is to characterize the role of the fluoxetine-resistant genes in serotonin downstream pathways that regulate developmental and physiological responses to environmental stresses. The fourth aim is to examine the genetic interactions between new fluoxetine-resistant genes and serotonin receptors. By testing serotonin deficient mutants, SERT-null mutants, serotonin receptor mutants, and fluoxetine resistant mutants, we will evaluate the comparability of serotonin, SSRIs and SERT on the function of serotonin receptors in regulating the synaptic activity of the other neurotransmitters, development, and stress physiology. PUBLIC HEALTH RELEVANCE: The systematic genetic dissection of SSRIs downstream targets in a model organism will reveal core components and principal functional pathways of the serotonergic system that exert conserved functions in disparate organisms and are important for the therapeutic effects of SSRIs. These functional pathways may also provide biochemical markers for subdividing a particular disease and reveal shared genetic vulnerability that might unify distinct disorders. These in turn will provide clues about the genetic basis underlying the pathogenesis and offer new windows for the discovery of novel therapeutics.